From a functional viewpoint, animal cells are roughly classified into parenchymal cells and non-parenchymal cells. Among these, parenchymal cells fulfill functions of tissues or organs. For instance, hepatocytes, i.e., parenchymal cells of liver, undertake the synthesis, decomposition and storage of various substances, and are therefore very important basic units for a living organism.
Thus, there have been proposed some kinds of system for the cultivation of hepatocytes by which to imitate in vitro the expression of the function of hepatocytes in a living organism. For example, R. Singhvi et al., U.S. Pat. No. 5,976,826, disclose a process to cultivate hepatocytes in a cytophilic regions which are separated by regions that are composed of hydrophilic and cytophobic substances, and a device therefor. In this device, individual cells are seeded on surface regions (generally 1-2,500 μm2, preferably 1-500 μm2) which are composed either of compounds which have hydrophobic surface or charged moieties (—COO−, —PO3H−) or of extracellular matrix proteins or the like. The cultivation which is mainly mentioned or intended in this U.S. Pat. No. 5,976,826 is regarded as so-called monolayer cultivation. On the other hand, there has also been tried three-dimensional architecture (three-dimensional growth pattern) with a view to improving the function of liver, e.g., enhancing the secretion of liver-enriched protein (M. Smalley et al. In Vitro Cell. Dev. Biol. Anim. (1999) 35, pp 22-32). In such three-dimensional culture methods, cell strains which have been derived from normal human liver are cultured on a gel of collagen type-I or of a special extracellular matrix.
There have further been provided devices by which to use cultured parenchymal or non-parenchymal cells conveniently in accordance with objectives such as toxicity test, and in which cultured cells are arrayed according to a certain pattern [e.g., Japanese Patent Application Laid-Open (Kokai) No. Hei 3 (1991)-7576 (a device for the control of cell-arrangement which device has cell-adhesive surface and non cell-adhesive surface), the above-mentioned U.S. Pat. No. 5,976,826 (the cultivation of hepatocytes), Japanese Patent No. 2973976 (the cultivation of endothelial cells), Japanese Patent Application Laid-Open (Kokai) No. Hei 7 (1995)-308186 (the cultivation of endothelial cells), etc.].
As mentioned above, it seems generally that the function of cultured cells which are given by three-dimensional architecture (three-dimensional growth pattern) is closer to that of hepatocytes in vivo than the function of cultured cells which are given by monolayer cultivation. Cultured cells of such a three-dimensional architecture system are liable to be peeled off the culture support, and, therefore, no three-dimensional structures have ever been realized on a patterned surface. Thus, three-dimensional architecture (three-dimensional growth pattern) is still unsatisfactory in the degree of expression of function and in the span of maintenance of function.
To begin with, organs of animals are constituted by tissues (groups of cells having similar function) of different properties. In most cases, cells which are units to constitute tissues keep their function through interaction between the same or different species of cells. Hence, from a viewpoint that heterotypic cellular interaction between parenchymal cells and adjacent non-parenchymal cells modifies cellular growth, migration and/or differentiation, S. N. Bhatia et al. set forth results of the study of the preservation of phenotype of hepatocyte which is given by the cocultivation of hepatocytes and fibroblasts which are non-parenchymal cells [S. N. Bhatia et al., The FASEB Journal, Vol. 13 (1999); 1883-1900]. In the monolayer of thus co-cultured hepatocytes which is enclosed by thus cultured non-parenchymal cells on the same plane, the intra-cellular albumin production ability, for instance, of hepatocytes which are located at a distance of three or four cells from the boundary between the monolayer of hepatocytes and the non-parenchymal cells decreases down to the same degree as that of purely cultured hepatocytes. In such a co-cultivation system, it is difficult for cells in an agglomerate of cultured hepatocytes to perform their liver-specific function stably.
Hence, the objective of the present invention is to provide a system of cultured animal cells by which to maintain specific function of parenchymal cells such as hepatocytes steadily and over a long period of time, and by which to keep micropattern stably when cells of the system are micropatterned.